The invention relates to optical imaging methods and systems, and more particularly to methods and systems for fluorescence imaging of a light scattering medium.
In vivo imaging of biological tissues facilitates early detection of disease, thereby providing an opportunity for reliable and pro-active diagnosis of diseased tissues. Fluorescence imaging is an example of a powerful non-invasive imaging technique that has been used in various applications in biological sciences. For example, fluorescence imaging is applied in fields such as genetic sequencing, biomedical diagnostics, and flow cytometry. Typically, fluorescence-imaging systems include a light source, which illuminates the subject to be imaged. The tissue inside the subject fluoresces either endogenously or exogenously in response to the excitation illumination, and the resulting emission is imaged to obtain information about the internal composition of the subject.
Fluorescence imaging is generally hampered by poor signal-to-background ratio (SBR) of fluorescent targets located within a subject. Much of this background noise is caused by reflection of the excitation light from the surface, and by strong fluorescence signals emitted from points near the surface of the subject. Different types of fluorescence imaging have been proposed to enhance SBR or contrast in fluorescence imaging. Some of these methods are fluorescence lifetime imaging, multi-spectral and hyper-spectral imaging. Fluorescence lifetime sensitive imaging (FLIM) differentiates various fluorescing species by their relative excited state lifetimes in a time- or frequency-resolved detection. For example, a fluorescence marker has a substantially different fluorescence decay rate than that of the tissue, it is possible to differentiate the two species by FLIM. FLIM reduces the effect of background auto-fluorescence, allowing greater contrast than conventional fluorescence imaging. However, in majority of cases, the difference of lifetime between many existing fluorescence markers and tissue is small, limiting the utility of FLIM.
Multi-spectral imaging has been demonstrated to be able to differentiate auto-fluorescence background and labeled markers for in-vivo applications. Typically, chemometric analysis is used to distinguish spectral signal that originates from the fluorescence marker and tissue auto-fluroescence background since they have distinct features. However, when the concentration of the bio-marker in the tissue is relatively low, unreliable results may be obtained. In addition, such an analysis may result in loss of a substantial amount of signal as only a narrow spectral region of the fluorescence signal is acquired for the analysis. Hence, such an analysis may not be ideal for real time applications.
Accordingly, there is a need for imaging systems and methods that can provide enhanced contrast in fluorescence imaging.